One of the fundamental components of morphogenesis is the ability of cells to segregate from each other to form new tissues or organs. The earliest example of this during vertebrate development is the formation of the primary germ layers. In Xenopus, each germ layer arises from a separate area of the spherical blastula. The ectoderm arises from animal cells, the mesoderm from marginal cells, and the endoderm from vegetal cells. Lineage analysis, the expression of regional markers, tissue recombination experiments, cell sorting experiments, and cell transplantation experiments all suggest that specification of the germ layers occurs during the mid- to late-blastula stages, before overt segregation of the germ layers. Vegetal cells inherit a maternally encoded transcription factor, VegT. This activates the expression of other transcription factors that initiate endoderm differentiation, and signaling molecules that induce the neighboring marginal zone cells to form mesoderm. The pathway leading to ectoderm formation is unknown. Preliminary data suggests that it is also initiated by mRNAs transcribed in the oocyte and inherited by the egg. We have found that a LIM homeodomain protein, XLIM5, already known to be expressed only in the ectoderm, is an immediate target of maternal transcription factor(s), and, when over expressed in early vegetal cells, causes their descendants to segregate to other germ layers. The goals of this project are to use this as a starting point to identify the pathway leading to ectoderm specification and the segregation of animal cells to form the ectoderm germ layer. We will identify the upstream maternal factors that activate XLIM5 expression, and the downstream genes which confer ectoderm-type adhesion on animal cells, and test their individual functions by simple gain- and loss-of function assays. We will also investigate the interaction between the ectoderm and endoderm pathways that leads to differential adhesion behaviour at the blastula stage. [unreadable] [unreadable]